title: Dependence of adenine production upon polyamine synthesis in cultured human lymphoblasts.
abstract: The exact source of de novo adenine produced by mammalian cells remain poorly understood, and this prompted the present study. Using a human lymphoblastoid cell line (WI-L2) deficient in adenine phosphoribosyltransferase (EC 2.4.2.7), we have quantitated the rate of adenine synthesis and the relative importance of the phosphorolysis of 5'-methylthioadenosine versus adenosine or 2'-deoxyadenosine in adenine generation. Dividing adenine phosphoribosyltransferase-deficient WI-L2 cells produced adenine at a rate of 0.27 nmol/mg protein/h. This represented approximately 10% of the rate of hypoxanthine production by WI-L2 cells deficient in hypoxanthine phosphoribosyltransferase (EC 2.4.2.8) but was equivalent to the rate of 5'-methylthioadenosine synthesis by human lymphoblastoid CCRF-CEM deficient in 5'-methylthioadenosine, phosphorylase (5'-methylthioadenosine: orthophosphate methylthioribosyltransferase). Up to 97% of adenine, but not hypoxanthine, synthesis was inhibited dose-dependently by the S-adenosylmethionine decarboxylase-inhibitor methylglyoxal bis(guanylhydrazone) and also by spermidine and spermine, but was enhanced by putrescine. The addition of 2-fluoroadenine, a potent competitive inhibitor of methylthioadenosine phosphorylase (Ki = 0.43 microM) to adenine phosphoribosyl-transferase-deficient cells resulted in a progressive accumulation of 5'-methylthioadenosine in the culture medium, and up to an 85% decrease in adenine production at non-toxic concentrations. These results show that de novo adenine synthesis by dividing human cells is considerable, and that 85-97% derives from the cleavage of 5'-methylthioadenosine and hence from polyamine synthesis.
mesh_terms: D000225:Adenine / Q000096:biosynthesis*; D000228:Adenine Phosphoribosyltransferase / Q000172:deficiency; D000241:Adenosine / Q000031:analogs & derivatives / Q000172:deficiency; D012437:Adenosylmethionine Decarboxylase / Q000037:antagonists & inhibitors; D002478:Cells, Cultured; D014644:Genetic Variation; D006801:Humans; D007041:Hypoxanthine Phosphoribosyltransferase / Q000172:deficiency; D007945:Leukemia, Lymphoid / Q000378:metabolism; D011073:Polyamines / Q000096:biosynthesis*; D011683:Purine-Nucleoside Phosphorylase / Q000172:deficiency; D011700:Putrescine / Q000096:biosynthesis; D013095:Spermidine / Q000096:biosynthesis; D013096:Spermine / Q000096:biosynthesis; D013154:Spleen / Q000378:metabolism; D013872:Thionucleosides / Q000172:deficiency
chemical_list: D011073:Polyamines; D013872:Thionucleosides; D013096:Spermine; D011683:Purine-Nucleoside Phosphorylase; C021784:5'-methylthioadenosine phosphorylase; D000228:Adenine Phosphoribosyltransferase; D007041:Hypoxanthine Phosphoribosyltransferase; D012437:Adenosylmethionine Decarboxylase; D000225:Adenine; D000241:Adenosine; D013095:Spermidine; D011700:Putrescine